Abstract
Hydrogen sulfide (H2S), generated in the osteoblasts predominantly via cystathionine-γ-lyase (CSE), is bone protective. Previous studies suggested that the onset of bone loss due to estrogen deficiency is associated to decreased levels of H2S and blunted gene expression of CSE. However, there are still a lot of unknowns on how H2S levels influence bone cells function. The present study aims to explore the mechanisms by which estrogen may regulate CSE expression, in particular the role of estrogen receptor alpha (ERα) in human osteoblasts (hOBs). Vertebral lamina derived hOBs were characterized and then assessed for CSE expression by western blot analysis in the presence or absence of ERα overexpression. Bioinformatic analysis, luciferase reporter assay and ChIP assay were performed to investigate ERα recruitment and activity on hCSE gene promoter.Three putative half Estrogen Responsive Elements (EREs) were identified in the hCSE core promoter and were found to participate in the ERα – mediated positive regulation of CSE expression. All osteoblast samples responded to ERα over-expression increasing the levels of CSE protein in a comparable manner. Notably, the ERα recruitment on the regulatory regions of the CSE promoter occurred predominantly in female hOBs than in male hOBs. The obtained results suggest that CSE/H2S system is in relation with estrogen signaling in bone in a gender specific manner.
Highlights
Cystathionine gamma-lyase (CSE) is the predominant hydrogen sulfide (H2S)-producing enzyme in mammalian cells other systems including that supported by cystathionine β-synthase (CBS) exist [1].The interest in the CSE enzyme and its regulation is growing, due to the recently described functions of H2S including the S-sulphydration of various target proteins [2]
To test the hypothesis that estrogen protects from bone loss [17] through CSE up-regulation in osteoblasts, we investigated whether the estrogen receptor alpha (ERα) directly participates in the regulation of CSE gene promoter activity and in the modulation of CSE expression in human male and female osteoblasts
Mature osteoblast phenotype was verified by analyzing the expression of the osteogenic markers including alkaline phosphatase (ALP), Runx2 transcription factor belonging to Runt-related factors family, osteopontin (OPN), and Collagen type I (COL1)
Summary
Cystathionine gamma-lyase (CSE) is the predominant hydrogen sulfide (H2S)-producing enzyme in mammalian cells other systems including that supported by cystathionine β-synthase (CBS) exist [1].The interest in the CSE enzyme and its regulation is growing, due to the recently described functions of H2S including the S-sulphydration of various target proteins [2]. The antioxidant and anti-inflammatory properties together with the cytoprotective effects of H2S seem to be so critical that abnormal H2S metabolism has been linked to several human pathologies, including autoimmune diseases, Alzheimer’s, hypertension, coronary heart disease, atherosclerosis, cataracts, pancreatitis, type 1 diabetes, osteoporosis and rheumatoid arthritis [3,4,5,6]. It is www.impactjournals.com/oncotarget urgent to understand the tissue-dependent control of its production. Glucose, heme oxygenase-1/carbon monoxide (HO-1/CO) system, and various CSE inhibitors suppress CSE activity and, H2S production [11, 12]
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